Cloning, expression and characterization of insulin-degrading enzyme from tomato (Solanum lycopersicum)

2008 ◽  
Vol 389 (1) ◽  
pp. 91-98 ◽  
Author(s):  
Yoann Huet ◽  
Jochen Strassner ◽  
Andreas Schaller
Keyword(s):  
Solanum Lycopersicum ◽  
Catalytic Efficiency ◽  
Biochemical Properties ◽  
Amyloid Peptide ◽  
Energy Status ◽  
Insulin Degrading Enzyme ◽  
Peptide Bonds ◽  
Degrading Enzyme ◽  
Β Amyloid Peptide

Abstract A cDNA encoding insulin-degrading enzyme (IDE) was cloned from tomato (Solanum lycopersicum) and expressed in Escherichia coli in N-terminal fusion with glutathione S-transferase. GST-SlIDE was characterized as a neutral thiol-dependent metallopeptidase with insulinase activity: the recombinant enzyme cleaved the oxidized insulin B chain at eight peptide bonds, six of which are also targets of human IDE. Despite a certain preference for proline in the vicinity of the cleavage site, synthetic peptides were cleaved at apparently stochastic positions indicating that SlIDE, similar to IDEs from other organisms, does not recognize any particular amino acid motif in the primary structure of its substrates. Under steady-state conditions, an apparent K m of 62±7 μm and a catalytic efficiency (k cat/K m) of 62±15 mm -1 s-1 were determined for Abz-SKRDPPKMQTDLY(NO3)-NH2 as the substrate. GST-SlIDE was effectively inhibited by ATP at physiological concentrations, suggesting regulation of its activity in response to the energy status of the cell. While mammalian and plant IDEs share many of their biochemical properties, this similarity does not extend to their function in vivo, because insulin and the β-amyloid peptide, well-established substrates of mammalian IDEs, as well as insulin-related signaling appear to be absent from plant systems.

scholarly journals Ablation of amyloid precursor protein increases insulin-degrading enzyme levels and activity in brain and peripheral tissues

2019 ◽  
Vol 316 (1) ◽  
pp. E106-E120 ◽  
Author(s):  
Joshua A. Kulas ◽  
Whitney F. Franklin ◽  
Nicholas A. Smith ◽  
Gunjan D. Manocha ◽  
Kendra L. Puig ◽  
...  
Keyword(s):  
Amyloid Precursor Protein ◽  
Ex Vivo ◽  
Precursor Protein ◽  
Amyloid Peptide ◽  
Type I ◽  
Insulin Degrading Enzyme ◽  
Peripheral Tissues ◽  
Degrading Enzyme ◽  
Bodies Of Evidence

The amyloid precursor protein (APP) is a type I transmembrane glycoprotein widely studied for its role as the source of β-amyloid peptide, accumulation of which is causal in at least some cases of Alzheimer’s disease (AD). APP is expressed ubiquitously and is involved in diverse biological processes. Growing bodies of evidence indicate connections between AD and somatic metabolic disorders related to type 2 diabetes, and App−/− mice show alterations in glycemic regulation. We find that App−/− mice have higher levels of insulin-degrading enzyme (IDE) mRNA, protein, and activity compared with wild-type controls. This regulation of IDE by APP was widespread across numerous tissues, including liver, skeletal muscle, and brain as well as cell types within neural tissue, including neurons, astrocytes, and microglia. RNA interference-mediated knockdown of APP in the SIM-A9 microglia cell line elevated IDE levels. Fasting levels of blood insulin were lower in App−/− than App+/+ mice, but the former showed a larger increase in response to glucose. These low basal levels may enhance peripheral insulin sensitivity, as App−/− mice failed to develop impairment of glucose tolerance on a high-fat, high-sucrose (“Western”) diet. Insulin levels and insulin signaling were also lower in the App−/− brain; synaptosomes prepared from App−/− hippocampus showed diminished insulin receptor phosphorylation compared with App+/+ mice when stimulated ex vivo. These findings represent a new molecular link connecting APP to metabolic homeostasis and demonstrate a novel role for APP as an upstream regulator of IDE in vivo.

In Vivo Aggregation of β-Amyloid Peptide Variants

2002 ◽  
Vol 71 (4) ◽  
pp. 1616-1625 ◽  
Author(s):  
David S. Fay ◽  
Amy Fluet ◽  
Carolyn J. Johnson ◽  
Christopher D. Link
Keyword(s):  
Amyloid Peptide ◽  
Β Amyloid ◽  
Β Amyloid Peptide

scholarly journals CALHM1 ion channel elicits amyloid-  clearance by insulin-degrading enzyme in cell lines and in vivo in the mouse brain

2015 ◽  
Vol 128 (13) ◽  
pp. 2330-2338 ◽  
Author(s):  
V. Vingtdeux ◽  
P. Chandakkar ◽  
H. Zhao ◽  
L. Blanc ◽  
S. Ruiz ◽  
...  
Keyword(s):  
Ion Channel ◽  
Cell Lines ◽  
Mouse Brain ◽  
Insulin Degrading Enzyme ◽  
Degrading Enzyme

scholarly journals Comparison of in vitro and in vivo activities associated with the G6PD allozyme polymorphism in Drosophila melanogaster.

Genetics ◽  
1990 ◽  
Vol 125 (4) ◽  
pp. 845-853
Author(s):  
W F Eanes ◽  
L Katona ◽  
M Longtine
Keyword(s):  
Protein Level ◽  
Catalytic Efficiency ◽  
Biochemical Properties ◽  
Activity Level ◽  
G6pd Activity ◽  
P Element ◽  
Allozyme Polymorphism ◽  
Low Activity

Abstract Earlier studies of the A and B allozymes at the G6pd locus show a differential ability of the genotypes to suppress the loss of viability associated with a low activity 6-phosphogluconate dehydrogenase mutation, 6Pgdlo1. This observation indicates a relatively lower activity for the A allozyme genotype, but it is not known if this level of suppression required a large difference in in vivo activity. To clarify this difference an analysis of the biochemical properties of the purified allozymes was carried out, as well as an analysis of the activity level associated with an original low activity P element-derived allele which had partially reverted and lost its suppression ability. G6PD activity and protein level were studied in 47 X chromosome lines from North America. The A genotype averages a 9% lower Vmax. From analysis of the correlation between G6PD activity and protein level it remains unclear whether the allozyme Vmax difference results from dissimilarity in protein level or kcat. At 25 degrees and physiological pH, comparative studies of the steady-state kinetics show the two purified allozyme variants differ significantly in their KM values for glucose-6-phosphate and NADP, and the K1 for NADPH. In aggregate these parameters predict the A genotype possesses a 20% lower in vitro catalytic efficiency. A partial revertant of a P element-derived low activity B variant, was shown to lose the ability to suppress 6Pgdlo1 low viability after acquiring only 60% of normal B activity. This last comparison shows the A genotype activity must be reduced in vivo by at least 40%.

scholarly journals Assessment of the Bioactivity of Antibodies against β-Amyloid Peptide in vitro and in vivo

2004 ◽  
Vol 1 (4-5) ◽  
pp. 160-167 ◽  
Author(s):  
M. Hasan Mohajeri ◽  
Meret N.M. Gaugler ◽  
Julia Martinez ◽  
Jay Tracy ◽  
Hong Li ◽  
...  
Keyword(s):  
Amyloid Peptide ◽  
Β Amyloid ◽  
Β Amyloid Peptide

scholarly journals Mutant presenilins specifically elevate the levels of the 42 residue β-amyloid peptide in vivo: evidence for augmentation of a 42-specific γ secretase

2003 ◽  
Vol 13 (2) ◽  
pp. 159-170 ◽  
Author(s):  
Joanna L. Jankowsky ◽  
Daniel J. Fadale ◽  
Jeffrey Anderson ◽  
Guilian M. Xu ◽  
Victoria Gonzales ◽  
...  
Keyword(s):  
Amyloid Peptide ◽  
Β Amyloid ◽  
Β Amyloid Peptide
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